Heparan sulfate proteoglycans (HSPGs) regulate numerous cell surface signaling events. They are extracellular modulators of signal transduction pathways during development and disease. HSPGs are cell-surface proteins that mainly consist of glycosylphosphatidylinositol (GPI)-anchored glypicans and transmembrane syndecans. Several HSPGs are currently being evaluated as potential targets for cancer therapy because of their relatively high expression in certain tumor types. In recent years, we have studied glypicans as a new class of cancer targets. Glypican-3 (GPC3) is a new therapeutic target in hepatocellular carcinoma (HCC), the most common form of primary liver cancers. We produced several antibodies targeting GPC3 either by hybridoma and phage display technologies. To isolate high affinity antibodies (e.g. YP7) to the native form of cell surface antigens such as GPC3, we developed a new high-throughput method combining functional cell binding screening by flow cytometry and conventional hybridoma technology [Phung et al., MAbs, PMID 22820551, 2012]. Furthermore, we have developed a new approach to humanize non-human antibodies (including mouse and rabbit antibodies) for clinical development [Zhang and Ho, Scientific Reports, 2016; Zhang and Ho, MAbs, 2017]. In addition, we used phage display technology to generate two human monoclonal antibodies (HN3 and HS20). HN3 is a human single-domain antibody that recognizes a novel functional site in the core protein of GPC3 and inhibits proliferation of HCC cells via blocking Wnt and Yap cancer signaling [Feng et al., PNAS, PMID: 23471984, 2013; Gao et al., Nature Communications, PMID: 25758784, 2015]. HS20 recognizes the heparan sulfate chains of GPC3. The human antibody disrupts the interaction of Wnt3a and GPC3 and inhibits Wnt/beta-catenin signaling [Gao et al., Hepatology, PMID: 24492943, 2014; Gao et al., PLoS One, PMID: 26332121, 2016; Gao et al., Scientific Reports, PMID: 27185050, 2016]. Our antibodies exhibit significant inhibition of HCC xenograft tumor growth in mice and show potential for use as therapeutic candidates. Furthermore, we found that GPC3 was efficiently internalized from the cell surface and that the HN3-PE38 immunotoxin brought the toxin into the cell, resulting in inhibition of protein synthesis. The immunotoxin caused regression of liver cancer in mice. Interestingly, Its novel mechanism involved both inhibition of cancer signaling (Wnt/Yap) and reduction in protein synthesis. Our strategy combining both antibody and toxin functions could be applicable generally to other immunotoxins and antibody-toxin/drug conjugates. To pursue clinical development of our anti-GPC3 immunotoxin for the treatment of liver cancer, we generated a new version of the anti-GPC3 immunotoxin (HN3-mPE24) and found that the second generation greatly reduced side effects and had better anti-tumor activity [Wang et al., Oncotarget, 2017]. In addition to the immunotoxin therapy, along with our collaborators, we used our anti-GPC3 antibodies to construct various clinical formats for targeted therapy of liver cancer including chimeric antigen receptor (CAR) T cell immunotherapy and photoimmunotherapy [Hanaoka et al. Mol Pharm, 2015; Hanaoka et al. Nanomedicine, 2015]. In addition to targeted therapies, our antibodies have been widely used as a research tool to analyze the role of glypicans in Wnt signaling and other important biological processes. We have identified the Wnt binding domain in heparan sulfate using our HS20 human antibody in collaboration with Dr. Jian Liu's lab in the University of North Carolina [Gao et al., Scientific Reports, 2016]. In addition to GPC3, our lab has been studying other glypican members (e.g. GPC2) as potential therapeutic targets in pediatric cancers (as presented in 2017 American Association for Cancer Research Annual Meeting in Washington DC, April 1-5, 2017). In the mesothelin project, we have used rabbit monoclonal antibody technology toidentify a panel of high affinity antibodies that bind novel sites in mesothelin. We have humanized one of the best candidates (YP218) for the treatment of mesothelioma and other mesothelin-positive cancers [Zhang et al., Scientific Reports, 2015; Zhang and Ho, MAbs, 2017].